We propose the development of a microfluidic real-time PCR microarray technology that is far beyond the capability of today's hybridization-based DNA microarrays. This new technology will enable a broad range of research and diagnostic applications that require DNA- and RNA-based assays which are massively parallel, quantitative, of high specificity and sensitivity, and possess a large dynamic range. We intend to apply the real-time PCR microarray as a standard nucleic acid detection engine to be used in research analytical systems, clinical diagnostic systems, public and home environmental monitors, field biothreat detectors, and many other instruments. Upon successful commercialization, the new real-time PCR microarrays will cost no more than today's regular DNA microarrays and will have the potential to eventually replace them as the next generation of DNA microarrays with fundamentally improved assay capabilities and added functionalities. The proposed microfluidic chip is made of either silicon/glass or plastic material and is a simple pressure-driven device, which involves a multitude of reaction cells interconnected by a uniquely designed fluidic network. The main advantages of the proposed new technology, as compared to the current 96 or 384 titer-plates based real-time PCR technology, include a significant reduction in the required amount of sample, a significant increase in the number of sequences simultaneously measured, a significant reduction in consumption of assay reagents, and a significant simplification in the operation of the assay. The goal of the Phase I project is to demonstrate the feasibility of performing parallel real-time PCR assays on a novel microfluidic array chip and lay the foundation for Phase II development. At the end of this proposed project, including Phase I and Phase II, we expect to have developed a prototype microfluidic array chip as a consumable product, a fabrication process for making the chip, and a prototype instrument for performing microfluidic real-time PCR assays at customer sites.